The primary goals of this application are to elucidate mechanisms of maturation of spermatozoa in the epididymis which will lead to an understanding of how fertility is regulated in the male. Maturation of sperm in the epididymis involves interactions among sperm, luminal fluid, and the epididymal epithelium (which is regulated by factors from the blood vascular system). In the experiments to be done in this application, investigation will be undertaken on proteins which are synthesized and secreted by the epididymal epithelium. Antibodies raised against these proteins cross-react with molecules on the sperm surface as well as with molecules that are derived from the testis. The first aim of this grant proposal is to isolate pure compounds from rete testis fluid, from the epididymis, and from sperm surfaces, which will then be studied extensively with respect to their amino acid sequences and the carbohydrate components of their oligosaccharide chains. The underlying theme of these experiments is to determine whether all of the molecules that cross-react immunologically are actually structurally the same, and whether they are the same as the Alpha-lactalbumin from rat milk. After deducing their structure, the molecules will be studied with respect to their ability to modify galactosyltransferase activity, which is a major function of Alpha-lactalbumin. The third aim in the project is related to studies on the synthesis and secretion (and control of these two functions) of immunologically cross-reactive molecules from the epididymis in culture. We have developed a perifusion culture system for epididymal tubules that will allow us to do studies on long-term cultures of epididymis, in which we know that epididymal epithelium and sperm are reasonably normal. Finally, using the well characterized monoclonal antibodies raised against the individual polypeptides of the epididymal Alpha-lactalbumin complex, experiments will be done to determine whether the molecules have any effect on sperm motility or fertilizing ability.